The Mechanism Of Control By Biological Agents To Appple Scar Skin Viroid And Its Effects In Fields

Apple is the main cultivated fruit tree in China.Apple is one of the mian agricultural production in China.Apple virus diseases are important diseases threatening the sustainable development of apple production.There is no effective chemical to control apple virus up to date.In this study,the new protein resistance inducer ATai Ling and Bacillus amyloliquefaciens agent were used to test for controlling Apple scar skin virus.In order to explore their possible mechanisms against apple virus,the expression levels of pathogenesis-related protein genes of apple callus were detected by qRT-PCR.The main results are as following:1.The apple fruit with mottle symptoms or Hualian were identified as the Apple scar skin viroid(ASSVd).From the apple mosaic leaves Apple necrotic mosaic virus(ApNMV)was detected,but no Apple mosaic virus(ApMV)was detected.The specific primers for Aple chlorotic leaf spot virus(ACLSV),Apple stem pitting virus(ASPV)and Apple stem grooving virus(ASGV)were designed,and the qRT-PCR detection system for amplifying these viruses at the same amplified system was established.The detection systems were specific,and with high applified quliaty.Its sensitivity of detection was 100 times highter than conventional RT-PCR.The correlation coefficient of the established standard curves of ACLSV,ASGV and ASPV were above 0.980,and the amplification efficiencies were between 90% and 105%.The results showed that the method was highly specific,sensitive,and efficient for these viruses.2.ATai Ling was used by irrigating to root and spraying on leaf.In fruit mature period,the control effect were calculated by diseased rate of fruit.The results showed that the control effect on Fuji varieties was 91.7%,92.3% on G20 Fuji,and 67.3% on Qinmi.It indicated that ATai Ling could effect on apple scar skin vivoid diseases in different varieties.3.Bacillus amyloliquefaciens DP-03 Agent was used by irrigating to root and spraying on leaf.The control effects were calculated.The results showed that the control effect of DP-03 on Fuji variety was 84.1%,77.0% on Qinmi.The result showed that Bacillus amyloliquefaciens agent could control apple scar skin viroid effectively.4.Apple callus tissues were treated by ATai Ling and the expression of pathogenesis-related protein genes of apple callus were detected by qRT-PCR,such as PR1 a,PR8,PR10 a,PR10b,PR10 c,PR10d and DEF1.The results showed that these genes were induced by ATai Ling.The maximum relative expression of PR1 a was 5.0 times,the PR8 could reach 41.2 times,the expression of PR10 a,PR10b,PR10 c and PR10 d were about 8 times.The DEF1 gene was slightly induced,and the maximum relative expression was 3.9 times.These results suggested that ATai Ling may increase resistance to pathogens by inducing pathogenesis-related proteins.5.After treated by Bacillus amyloliquefaciens Agent DP-03,apple callus tissues were used to detect the expression of pathogenesis-related protein genes such as PR1 a,PR8,PR10 a,PR10b,PR10 c,PR10d and DEF1 by qRT-PCR.The results showed that Bacillus amyloliquefaciens could induce PR8 gene,PR10 a gene,PR10 b gene,PR10 c gene,PR10 d gene up-regulated,and could not induce PR1 a gene and DEF1 gene.The up-regulated expression of PR8 gene was the highest,and the maximum relative expression was 17.6times.The maximum relative expression of PR10 a gene,PR10 b gene,PR10 c gene and PR10 d gene were 10.6 times,9.7 times,7.6 times and 8.8 times.The results indicated that Bacillus amyloliquefaciens may increase the resistance of apple tree by inducing pathogenesis-related proteins.